The overall objective of these studies is to examine the alterations in cellular growth control that occur during neoplastic transformation of rat mammary epithelial (RME) cells. The working hypothesis is that normal cells have a finite proliferative lifespan and that an important alteration in cellular growth potential occurs during cancer development which results in the acquisition of an enhanced, perhaps indefinite, proliferative lifespan. Acquisition of this enhanced proliferative potential is necessary but insufficient for complete neoplastic transformation and therefore represents a truly preneoplastic phenotype. The experiments will employ a newly developed culture system in which normal RME cells undergo 15 to 20 population doublings over 4 to 5 in vitro passages before exhibiting signs of senescence. This culture system will be used; a) to examine the hormone and growth factor requirements that induce long term proliferation of normal RME cells in vitro, b) to isolate populations of preneoplastic cells from mammary tissues of carcinogen treated rats and populations of neoplastic cells from mammary carcinomas based on their enhanced proliferative lifespan, 3) to examine the hormone and growth factor requirements of preneoplastic and neoplastic RME cells relative to those of normal RME cells in vitro. These studies address the mechanisms of the alterations in growth control that occur in RME cells during the stepwise acquisition of enhanced proliferative potential and neoplastic potential.